Two pairs of bioactive cyclohexene alkaloid enantiomers from the roots of Piper nigrum.

Two pairs of cyclohexene amide alkaloid enantiomers were obtained from the root of Piper nigrum. Their plane structures were established by NMR and HRESIMS spectra. The absolute configurations of 1a/1b and 2a/2b were determined by the comparison between the experimental and calculated electronic circular dichroism (ECD) spectra. All identified compounds were tested for inhibitory effects on acetylcholinesterase (AChE) in vitro. Notably, compounds 1b and 2b showed strong inhibitory effects on AChE and the interaction between proteins and compounds was discussed by molecular docking studies.

Toxic Effects of Arsenic on Four Freshwater Aquatic Species and Its Transformation Metabolism in Crucian Carp (Carassius auratus).

Inorganic arsenic is a well-known carcinogen that is much more toxic than its organic counterpart. While much is known about the accumulation and transformation of arsenic in marine organisms, little is known regarding these processes in freshwater aquatic species. In this study, the acute toxicity and toxicological effects of inorganic arsenic on four freshwater organisms (Cyprinus carpio, Misgurnus anguillicaudatus, Pseudorasbora parva, Eriocheir sinensis) commonly found in rice-fish farming systems were investigated. The organisms exhibited different levels of sensitivity to inorganic arsenic, with crustaceans being more sensitive than fish. Fish were found to be more tolerant to As(V) than As(III). The study also investigated the accumulation, transformation, and release of inorganic arsenic in crucian carp, an omnivorous species with high environmental tolerance. The fish accumulated As(III) rapidly in various tissues, and were able to transport it to other tissues through gills, intestines, and skin. The accumulated As(III) was converted into less toxic forms, such as monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA), via methylation. The fish also converted As(III) into arsenate (AsV) via enzymatic and oxidative reactions. After the transferal to clean water, the forms of arsenic in the various tissues decreased rapidly, but the rates of excretion of the four forms of arsenic were not the same among the different tissues. Our results suggest that crucian carp can reduce the environmental toxicity of As(III) at certain concentrations by transforming it into less toxic forms within their bodies.

Expanded Application of Piper nigrum: Guided Isolation of Alkaloids with Inhibitory Activities of AChE/BuChE and Aß Aggregation.

Piper nigrum is a popular crop that can be used as seasoning or as an additive but its active ingredients also have an effect on the nervous system. Nineteen new amide alkaloids (1a/1b, 2-5, 6a/6b, 7, 8a/8b, 9, 10a/10b, 11a-11b, 12-14) were isolated from P. nigrum, guided by inhibitory activity of AChE and LC-MS/MS based on GNPS. The configurations were determined by extensive spectral analysis, Bulkiness rule, and NMR calculations. The inhibitory activities of AChE/BuChE and Aß aggregation were tested, and the results showed compounds 2, 7, and 12 had significant inhibitory activities. These components were identified in the crude fraction and their relative quantities were tested, which suggested that compound 2 was the index component in the active site from P. nigrum.

Utilization of the By-Product of Corn: Guided Identification of Bioactive Terpenoids from Stigma Maydis (Corn Silk).

Stigma maydis (corn silk) (S. maydis) is a food-based by-product of maize and possesses great nutritional and pharmaceutical value. This study aimed to explore bioactive components from S. maydis. By the guidance of bioactivity-guided approach and Global Natural Products Social (GNPS) molecular networking, 12 terpenoids were discovered from S. maydis. The structures of 11 undescribed compounds (1-11) were determined by detailed spectroscopic analyses, single-crystal X-ray diffraction analysis, specific rotation calculations, electronic circular dichroism (ECD) calculations, and NMR calculations. The neuroprotective and acetylcholinesterase (AChE) inhibitory effects of 1-12 were examined, and most of them showed significant or moderate activities. The underlying neuroprotective mechanism of 4 and 5 was revealed by Hoechst 33258, AO-EB, and JC-1 staining assays. This work illustrated the potential of S. maydis as a prospective natural source of bioactive compounds in food and pharmaceutical industries.

Identification and prevalence investigation of cyathocotylid trematode (Trematoda: Digenea: Cyathocotylidae) in Carassius auratus in Wuhu, China.

The family Cyathocotylidae trematode is a world-widely distributed parasite whose adults are mainly found in fish, reptiles, birds, and mammals in both freshwater and marine environments. However, little is known of the prevalence of these trematode in China. For the first time, we found Cyathocotylidae trematode in the Wuhu area. Therefore, we investigated the prevalence of metacercariae in Carassius auratus from Zhang Lake, Kui Lake, Qingyi River, and Yangtze River (Wuhu Section) in Wuhu area. A total of 392 one-year-old C. auratus were tested in January, April, July, and October 2019, respectively. After the fish were euthanized, the back, chest, and tail muscles were used for preliminary screening for the presence of metacercariae by direct compression method. Metacercariae were isolated by the artificial digestion method and then morphologically and molecularly identified by cox1 (642 bp) and ITS2 (418 bp) sequences amplification by PCR. Based on morphological features and sequence analysis, they were identified as cyathocotylid metacercariae. The four water sources have different degrees of prevalence throughout the year, the prevalence ranges from 28.45% (29/102) in Yangtze River to 50.55% (46/91) in Zhang Lake, with an average prevalence of 40.56% (159/392). There were also differences in the prevalence in different months: The highest occurred in July, which was 81.63% (80/98), while the lowest occurred in January, which was 8.89% (8/90). This study can provide basic data for the prevention and control of cyathocotylid trematode in this area.

HSQC-based small molecule accurate recognition technology discovery of diverse cytotoxic sesquiterpenoids from Elephantopus tomentosus L. and structural revision of molephantins A and B.

Elephantopus tomentosus L. is a perennial herb taxonomically belonging to the family Asteraceae, which has been used as a folk medicine for the treatment of hepatobiliary diseases. Sesquiterpenoids from this plant have broad biological activities, including anti-tumor, anti-inflammatory, and antibacterial effects. In this study, fifteen structurally diverse sesquiterpenoids comprised 11 germacrane-type and 4 eudesmane-type sesquiterpenoids were prioritized to isolated from Elephantopus tomentosus L. based on the HSQC-based Small Molecule Accurate Recognition Technology (SMART) strategy. Among them, ten sesquiterpenoids were previously unreported, and their structures were elucidated by spectroscopic data, computational methods, single-crystal X-ray diffraction crystallographic data or electronic circular dichroism calculations. In addition, the structures of two known sesquiterpenoids, molephantin A and B, which were reported to possess E-geometry for the Δ1(10) double bond, were revised by reanalyzing their spectroscopic and X-ray crystallographic data. Some sesquiterpenoids exhibited significant cytotoxic activities against Hep3B and HepG2 cell lines.

Two new iridoids and triterpenoid analogues from the leaves of Viburnum chingii and their anti-acetylcholinesterase activity.

Two undescribed split-ring iridoids (1-2) with six known triterpenes (3-8) and one steride (9) were isolated from the Viburnum chingii. Compound 2 possessed an unprecedented split-ring iridoid skeleton formed by electrocyclic reaction and split ring. The structures and absolute configurations of the new iridoids were established by NMR, HRESIMS, and ECD calculations. All the isolated compounds were tested for AChE inhibitory activity. Biologically, 1, 2, 3, 4, and 7 displayed significant AChE effects compared to the positive control donepezil, and have also been subjected to molecular docking studies.

Novel Bisamide Alkaloids Enantiomers from Pepper Roots (Piper nigrum L.) with Acetylcholinesterase Inhibitory and Anti-Neuroinflammatory Effects.

The roots of Piper nigrum L., a seasoning for cooking various types of broths, are renowned for their high nutritional content and potential medicinal benefits. In this study, nine pairs of novel cyclohexene-type bisamide alkaloids (1a/1b-9a/9b) were isolated from the pepper roots using molecular network analysis strategies. Their structures were determined by extensive spectroscopic data, electronic circular dichroism (ECD) calculations, and X-ray diffraction analyses. Using an intermolecular Diels-Alder reaction, a strategy for the synthesis of bisamide alkaloids from different monomeric amide alkaloids was developed. Furthermore, these compounds were chirally separated for the first time, and compounds 3a and 5a/5b showed significant anti-neuroinflammation effects in the models of lipopolysaccharide(LPS)-induced BV2 microglial cells. Meanwhile, compounds 6b and 7a displayed concentration-dependent inhibitory activities against acetylcholinesterase with IC50 values of 6.05 ± 1.10 and 3.81 ± 0.10 µM, respectively. These findings confirmed that these bisamide alkaloids could be applied in functional food formulations and pharmaceutical products as well as facilitate the further development and usage of pepper roots.

Highly Oxidized Germacranolides from Elephantopus tomentosus and the Configurational Revision of Some Previously Reported Analogues.

Highly oxidized germacranolides are mainly found in the genus Elephantopus, contain a characteristic ten-membered molecular core that is highly flexible, and exhibit potential cytotoxic properties. However, their configurations were assigned ambiguously in previous reports due to spectroscopic observation of macrocyclic systems. Herein, 17 highly oxidized germacranolides, including 12 new germacranolides (1-12), were isolated from Elephantopus tomentosus. Their structures were characterized by spectroscopic data analysis combined with X-ray crystallography and ECD calculations, and it was possible to propose configurational revisions of five previously reported analogues (13-17). Cytotoxic activities for 1-17 against two hepatocellular carcinoma cell lines (HepG2 and Hep3B) were tested, and compounds 1-10 and 13-16 generated IC50 values of 2.2-9.8 µM. Furthermore, the observed cytotoxic activity of 1 was determined as being mediated by inducing the apoptosis of HepG2 and Hep3B cells via mitochondrial dysfunction.

Chemical constituents from Daphne giraldii and their cytotoxicities and inhibitory activities against acetylcholinesterase.

Seven triterpenoids (1-7), two prenylated coumarins (8 and 9), and one diphenylpropane (10), including five previously undescribed compounds (1-3, 8, and 10), were obtained from the stem and root barks of Daphne giraldii. The structures and absolute configurations of the new triterpenoids were established by NMR, HRESIMS, ECD calculations, and single-crystal X-ray diffraction analysis. All identified compounds were tested for cytotoxicities (human tumour cell line Hep3B) and inhibitory effects on AChE in vitro. Notably, prenylated coumarins (8 and 9) exhibited moderate cytotoxic activities and 3-hydroxy-substituted triterpenoids (2 and 4) showed mild inhibitory effects on AChE. Furthermore, compounds 2 and 4 have also been subjected to molecular docking studies to investigate the inhibitory mechanism.

Insight into Isolation and Characterization of Phenolic Compounds from Hawthorn (Crataegus pinnatifida Bge.) with Antioxidant, Anti-Acetylcholinesterase, and Neuroprotective Activities.

Recent epidemiologic studies have demonstrated a link between the consumption of daily functional fruits rich in phenols and the prevention of disease for neurodegenerative disorders. Hawthorn products are derived from the functional fruit hawthorn, which is rich in phenols and has been used around the world for centuries. In order to explore the phenolic components in hawthorn, the investigation of the ethanol extract led to the separation of five new phenol compounds (1a/1b, 2-4), including one pair of enantiomers (1a/1b), along with seven disclosed analogs (5-11). Their structures were elucidated based on extensive spectroscopic analyses and electronic circular dichroism (ECD). The compounds (1-11) were tested for antioxidant activities by 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonicacid) (ABTS), and ferric reducing antioxidant power (FRAP) methods. Apart from that, monomeric compounds 2, 4, and 6 exhibited more potent protective capabilities against H2O2 (hydrogen peroxide)-induced SH-SY5Y cells. Meanwhile, electronic analyses were performed using the highest occupied molecular orbital (HOMO), and the lowest unoccupied molecular orbital (LUMO) to analyze compounds 2, 4, and 6. Furthermore, compounds (1-11) measured acetylcholinesterase (AChE) inhibitory activities, and 2, 4, and 6 possessed greater AChE inhibitory activity than donepezil. At the same time, molecular docking was used to investigate the possible mechanism of the interaction between active compounds (2, 4, and 6) and AChE.

Chemical structures and anti-tyrosinase activity of the constituents from Elephantopus scaber L.

Four undescribed compounds including one germacrane-type sesquiterpene lactones (1), alkaloid (2) along with two neolignans (3-4) were isolated from Elephantopus scaber L. Their structures and absolute configurations were elucidated unambiguously by means of 1D and 2D NMR spectroscopic data analysis, and quantum chemical electronic circular dichroism calculations, as well as single-crystal X-ray crystallography. Their anti-tyrosinase activities have been evaluated in vitro and compound 2 exhibited significant inhibitory activity. Furthermore, molecular docking was performed to study the interaction patterns between 2 and the tyrosinase.

Coumarins from Sarcandra glabra (Thunb.) Nakai and Acetylcholinesterase Inhibiting Activity.

Nine coumarins including a pair of new enantiomers (1a/1b) and seven known compounds (2-8) were isolated from Sarcandra glabra (Thunb.) Nakai. Among them, compounds 1a and 1b were naturally occurring coumarin-phenylpropanoid conjugate enantiomers. Their structures were identified by NMR and ECD calculations. Compounds 1-8 were tested for acetylcholinesterase (AchE) inhibiting activity. The results of the enzymology experiment showed that compound 3 demonstrated obvious AchE inhibitory activity which showed the IC50 value of 1.982±0.003 µM, and the binding sites were predicted by molecular docking.

Application of solid-phase extraction: High-resolution mass spectrometry analysis strategy in the characterization and quantification of amide herbicides in aquatic products.

In this study, we developed a new method combining thin-layer ultrasonic extraction, efficient SPE purification, ultra-performance liquid chromatography separation, and high-resolution mass spectrometry characterization for seven amide herbicides in fishery products. In sample preparation, to rapidly increase the contact area between the fish meat and the extractant, the fish meat was smeared on a glass slide. This process resulted in quickly reaching the extraction equilibrium and relatively high extraction efficiency. In data analysis, a strategy for characterization and qualitative analysis was constructed by analyzing the fragmentation of amide herbicides using product ion scans. Isomeric pretilachlor and butachlor were separated chromatographically, while the coeluting isomers, alachlor and acetochlor, could be separated by differences in the fragmentation of their selected precursor ions. This method overcame the challenge of poor dispersion in the extractant caused by the high viscosity of fish meat, and the challenge of separation and characterization for isomers. Compared with other methods, the extraction efficiency was improved and the amide herbicides in aquatic products was characterized and quantified rapidly and accurately. Moreover, the qualitative information was much greater and provided an additional strategy for analytes identification. This rapid and accurate method will benefit workers involved in monitoring fishery.

Selective magnetic solid-phase extraction of amide herbicides from fish samples coupled with ultra-high-performance liquid chromatography with tandem mass spectrometry.

An efficient magnetic dummy template molecularly imprinted polymer nanocomposite was prepared using multi-walled carbon nanotubes as a support and metolachlor deschloro as a dummy template. The obtained nanocomposites were characterized using Fourier transform infrared spectroscopy, vibrating sample magnetometry, scanning electron microscopy, and transmission electron microscopy. The adsorption performance of the obtained nanocomposites was evaluated through binding experiments, including static adsorption, kinetic adsorption, and selective recognition studies. The obtained nanocomposites were successfully applied as selective sorbents for the magnetic solid-phase extraction of seven amide herbicides (alachlor, acetochlor, pretilachlor, butachlor, metolachlor, diethatyl ethyl, and dimethachlor) coupled with liquid chromatography-tandem mass spectrometry from fish samples. Under the optimized conditions, the limit of detection was 0.01-0.1 µg/kg. The obtained recoveries of the amide herbicides from the fish samples were in the range of 88.0 to 102.1% with a relative standard deviation of less than 7.5%. This method, which eliminated the effect of template leakage on qualitative and quantitative analysis was found to be superior to the methods reported in the literature. The results indicated that it could be successfully applied to analyze amide herbicides in fish samples with satisfactory recoveries.

Combined effects of perchlorate and hexavalent chromium on the survival, growth and reproduction of Daphnia carinata.

Perchlorate and hexavalent chromium (Cr(VI)) are common cocontaminants in aquatic environments due to their high water solubility, stability, mobility, and some coapplications. However, few studies have investigated their combined toxicity to organisms. In this work, we studied the acute and chronic toxicities of perchlorate and Cr(VI), alone and in combination, with survival, growth, and reproduction as endpoints using Daphnia carinata as a model organism. For a single contaminant, Cr(VI) was found to be more toxic than perchlorate to D. carinata not only in terms of survival but also in terms of growth and reproduction. In regard to the combined pattern, the interactive effects on survival, growth, and reproduction were mainly additivity, antagonism, and synergism, respectively, suggesting that the interactive response of perchlorate and Cr(VI) is endpoint-specific. Due to significant synergism, over 21 days of observation, the inhibition of 0.1 mg/L perchlorate and 0.2 mg/L Cr(VI) on cumulative offspring per female in the first seven broods reached 63.9 ± 3.6%, suggesting that long-term exposure to perchlorate and Cr(VI) at environmentally relevant concentrations may affect D. carinata reproduction in the natural environment. Our results will be significant for understanding the complicated combined toxicity of perchlorate and Cr to aquatic organisms.

Loss of 4.1N in epithelial ovarian cancer results in EMT and matrix-detached cell death resistance.

Epithelial ovarian cancer (EOC) is one of the leading causes of death from gynecologic cancers and peritoneal dissemination is the major cause of death in patients with EOC. Although the loss of 4.1N is associated with increased risk of malignancy, its association with EOC remains unclear. To explore the underlying mechanism of the loss of 4.1N in constitutive activation of epithelial-mesenchymal transition (EMT) and matrix-detached cell death resistance, we investigated samples from 268 formalin-fixed EOC tissues and performed various in vitro and in vivo assays. We report that the loss of 4.1N correlated with progress in clinical stage, as well as poor survival in EOC patients. The loss of 4.1N induces EMT in adherent EOC cells and its expression inhibits anoikis resistance and EMT by directly binding and accelerating the degradation of 14-3-3 in suspension EOC cells. Furthermore, the loss of 4.1N could increase the rate of entosis, which aggravates cell death resistance in suspension EOC cells. Moreover, xenograft tumors in nude mice also show that the loss of 4.1N can aggravate peritoneal dissemination of EOC cells. Single-agent and combination therapy with a ROCK inhibitor and a 14-3-3 antagonist can reduce tumor spread to varying degrees. Our results not only define the vital role of 4.1N loss in inducing EMT, anoikis resistance, and entosis-induced cell death resistance in EOC, but also suggest that individual or combined application of 4.1N, 14-3-3 antagonists, and entosis inhibitors may be a promising therapeutic approach for the treatment of EOC.

Mass loss and nutrient release during the decomposition of sixteen types of plant litter with contrasting quality under three precipitation regimes.

Mass loss and nutrient release during litter decomposition drive biogeochemical cycling in terrestrial ecosystems. However, the relationship between the litter decomposition process and the decomposition stage, precipitation, and litter quality has rarely been addressed, precluding our understanding of how litter decomposition regulates nutrient cycling in various ecosystems and their responses to climate change. In this study, we measured mass loss as well as carbon and nutrient releases during the decomposition of 16 types of leaf litter under three precipitation treatments over 12 months in a common garden experiment (i.e., using standardized soil and climatic conditions). Sixteen types of leaves were divided into three functional groups (evergreen, deciduous, and herbaceous). The objectives were to understand the effects of decomposition stages and precipitation regimes on litter decomposition and to examine the relationship between this effect and chemical properties. The mass loss and release of nitrogen and potassium were significantly higher in the 6- to 12-month stage of decomposition (high temperature and humidity) than in the 0- to 6-month stage. Phosphorus was relatively enriched in evergreen leaves after 6 months of decomposition. The rates of mass loss and nutrient release were significantly greater in herbaceous than in deciduous and evergreen leaves. Increasing precipitation from 400 to 800 mm accelerated mass loss and potassium release but decreased phosphorus release in the 0- to 6-month stage of decomposition. These results highlighted the contribution to and complexity of litter chemical properties in litter decomposition.

High-throughput metabolomics enables metabolite biomarkers and metabolic mechanism discovery of fish in response to alkalinity stress.

High throughput mass spectrometry (MS)-based metabolomics is a popular platform for small molecule metabolites analyses that are widely used for detecting biomarkers in the research field of environmental assessment. Crucian carp (Carassius carassius, CC) is an economically and ecologically important fish in Asia. It can adapt to extremely high alkalinity, providing us with valuable material to understand the adaptation mechanism for extreme environmental stress. However, the information on the metabolite biomarkers and metabolic mechanisms of CC exposed to alkaline stress is not entirely clear. We applied high-throughput UPLC-Q-TOF/MS combined with chemometrics to identify changes in the metabolome of CC exposed to different concentrations of alkalinity for long term effects. Metabolic differences among alkalinity-treated groups were identified by multivariate statistical analysis. Further, 7 differential metabolites were found after exposure to alkaline conditions. In total, 23 metabolic pathways of these differential metabolites were significantly affected. Alkalinity exposure resulted in widespread change in metabolic profiles in the plasma with disruptions in the phenylalanine metabolism, glycine, serine and threonine metabolism, pyruvate metabolism, tyrosine metabolism, etc. The integrated pathway analysis of the associated metabolites showed that tRNA charging, l-cysteine degradation II, superpathway of methionine degradation, l-serine degradation, tyrosine biosynthesis IV, etc. appear to be the most significantly represented functional categories. Overall, this study demonstrated that metabolic changes in CC played a role in adaptation to the highly alkaline environmental stress.